Simultaneous targeted and discovery-driven clinical proteotyping using hybrid-PRM/DIA.

BACKGROUND: Clinical samples are irreplaceable, and their transformation into searchable and reusable digital biobanks is critical for conducting statistically empowered retrospective and integrative research studies. Currently, mainly data-independent acquisition strategies are employed to digitize clinical sample cohorts comprehensively. However, the sensitivity of DIA is limited, which is why selected marker candidates are often additionally measured targeted by parallel reaction monitoring. METHODS: Here, we applied the recently co-developed hybrid-PRM/DIA technology as a new intelligent data acquisition strategy that allows for the comprehensive digitization of rare clinical samples at the proteotype level. Hybrid-PRM/DIA enables enhanced measurement sensitivity for a specific set of analytes of current clinical interest by the intelligent triggering of multiplexed parallel reaction monitoring (MSxPRM) in combination with the discovery-driven digitization of the clinical biospecimen using DIA. Heavy-labeled reference peptides were utilized as triggers for MSxPRM and monitoring of endogenous peptides. RESULTS: We first evaluated hybrid-PRM/DIA in a clinical context on a pool of 185 selected proteotypic peptides for tumor-associated antigens derived from 64 annotated human protein groups. We demonstrated improved reproducibility and sensitivity for the detection of endogenous peptides, even at lower concentrations near the detection limit. Up to 179 MSxPRM scans were shown not to affect the overall DIA performance. Next, we applied hybrid-PRM/DIA for the integrated digitization of biobanked melanoma samples using a set of 30 AQUA peptides against 28 biomarker candidates with relevance in molecular tumor board evaluations of melanoma patients. Within the DIA-detected approximately 6500 protein groups, the selected marker candidates such as UFO, CDK4, NF1, and PMEL could be monitored consistently and quantitatively using MSxPRM scans, providing additional confidence for supporting future clinical decision-making. CONCLUSIONS: Combining PRM and DIA measurements provides a new strategy for the sensitive and reproducible detection of protein markers from patients currently being discussed in molecular tumor boards in combination with the opportunity to discover new biomarker candidates.

Phosphorus-Water Interaction Drives Active Center Evolution into the Water-Adaptive Structure in the High-Humidity NH3-SCR Reaction.

The impact of water on catalyst activity remains inconclusive due to its dependence on the specific reaction environment. To maximize the exploitation of water's promoting effect, we employed ammonia selective catalytic reduction (NH3-SCR) as a probe reaction and proposed a phosphorus modification strategy for Cu-ZSM-5 catalysts. The objective of this approach was to construct water-adaptive microstructures through directional arrangement. To investigate the effect of phosphorus on the transformation of framework copper sites in humid environments, we conducted comprehensive characterizations and density functional theory calculation. Results reveal that water molecules cleave the oxygen bridges between phosphorus oxide and copper, leading to the formation of active isolated [Cu(OH)]+ groups and phosphate. The phosphate species weaken the interaction between exchanged Cu2+ groups and the zeolite framework, leading to the generation of highly migratory hydrated Cu2+ species. This work will potentially guide the rational design of water-adaptive catalysts for gas pollution abatement in a humid environment.

Bioaccumulation and Male Reproductive Toxicity of the New Brominated Flame Retardant Tetrabromobisphenol A-Bis(2,3-dibromo-2-methylpropyl ether) in Comparison with Hexabromocyclododecane.

Tetrabromobisphenol A-bis(2,3-dibromo-2-methylpropyl ether) (TBBPA-DBMPE) has come into use as an alternative to hexabromocyclododecane (HBCD), but it is unclear whether TBBPA-DBMPE has less hazard than HBCD. Here, we compared the bioaccumulation and male reproductive toxicity between TBBPA-DBMPE and HBCD in mice following long-term oral exposure after birth. We found that the concentrations of TBBPA-DBMPE in livers significantly increased with time, exhibiting a bioaccumulation potency not substantially different from HBCD. Lactational exposure to 1000 µg/kg/d TBBPA-DBMPE as well as 50 µg/kg/d HBCD inhibited testis development in suckling pups, and extended exposure up to adulthood resulted in significant molecular and cellular alterations in testes, with slighter effects of 50 µg/kg/d TBBPA-DBMPE. When exposure was extended to 8 month age, severe reproductive impairments including reduced sperm count, increased abnormal sperm, and subfertility occurred in all treated animals, although 50 µg/kg/d TBBPA-DBMPE exerted lower effects than 50 µg/kg/d HBCD. Altogether, all data led us to conclude that TBBPA-DBMPE exerted weaker male reproductive toxicity than HBCD at the same doses but exhibited bioaccumulation potential roughly equivalent to HBCD. Our study fills the data gap regarding the bioaccumulation and toxicity of TBBPA-DBMPE and raises concerns about its use as an alternative to HBCD.

Growth performance, nutrient digestibility, fecal microbial diversity and volatile fatty acid, and blood biochemical indices of suckling donkeys fed diets supplemented with multienzymes.

BACKGROUND: As the foal grows, the amount of breast milk produced by the donkey decreases. In such cases, early supplemental feeding is particularly important to meet the growth needs of the foal. Foals have an incompletely developed gastrointestinal tract with a homogenous microbiota and produce insufficient amounts of digestive enzymes, which limit their ability to digest and utilize forage. Improving the utilization of early supplemental feeds, promoting gastrointestinal tract development, and enriching microbial diversity are the hotspots of rapid growth research in dairy foals. Plant-based feeds usually contain non-starch polysaccharides (NSPs), including cellulose, xylan, mannan, and glucan, which hinder nutrient digestion and absorption. In addition, proteins and starch (both biomolecules) form a composite system mainly through non-covalent interactions. The proteins wrap around the surface of starch granules and act as a physical obstacle, thereby inhibiting water absorption and expansion of starch and decreasing the enzyme's catalytic effect on starch. Glyanase, ß-mannanase, ß-glucanase, cellulase, protease, and amylase added to cereal diets can alleviate the adverse effects of NSPs. The current study determined the effects of adding multienzymes (glyanase, ß-mannanase, ß-glucanase, cellulase, protease, and amylase) to the diet of 2-month-old suckling donkeys on their growth performance, apparent nutrient digestibility, fecal volatile fatty acid (VFA) and pH, fecal bacterial composition, and blood biochemical indices. RESULTS: On day 120 of the trial, fecal samples were collected from the rectum of donkeys for determining bacterial diversity, VFA content, and pH. Moreover, fresh fecal samples were collected from each donkey on days 110 and 115 to determine apparent digestibility. The multienzymes supplementations did not affect growth performance and apparent nutrient digestibility in the donkeys; however, they tended to increase total height gain (P = 0.0544). At the end of the study, the multienzymes supplementations increased (P < 0.05) the Observed species, ACE, Chao1, and Shannon indices by 10.56%, 10.47%, 10.49%, and 5.01%, respectively. The multienzymes supplementations also increased (P < 0.05) the abundance of Firmicutes, Oscillospiraceae, Lachnospiraceae, Christensenellaceae, Christensenellaceae_R-7_group, and Streptococcus in feces, whereas decreased (P = 0.0086) the abundance of Proteobacteria. CONCLUSIONS: Multienzymes supplementations added to a basal diet for suckling donkeys can increase fecal microbial diversity and abundance.

Extended exposure to tetrabromobisphenol A-bis(2,3-dibromopropyl ether) leads to subfertility in male mice at the late reproductive age.

Tetrabromobisphenol A-bis(2,3-dibromopropyl ether) (TBBPA-BDBPE), a commonly used brominated flame retardant as a decabromodiphenyl ether substitute, has been detected in various environmental compartments, but its health hazards remain largely unknown. Our recent study showed that low-dose exposure of male mice to TBBPA-BDBPE from postnatal day (PND) 0 to 56 caused remarkable damage to the microtubule skeleton in Sertoli cells and the blood-testis barrier (BTB) but exerted little effect on conventional reproductive endpoints in adulthood. To investigate whether TBBPA-BDBPE may cause severe reproductive impairments at late reproductive age, here, we extended exposure of historically administrated male mice to 8-month age and allowed them to mate with non-treated females for the evaluation of fertility, followed by a general examination for the reproductive system. As expected, we found that 8-month exposure to 50 µg/kg/d as well as 1000 µg/kg/d TBBPA-BDBPE caused severe damage to the reproductive system, including reduced sperm counts, increased sperm abnormality, histological alterations of testes. Moreover, microtubule damage and BTB-related impairment were still observed following 8-month exposure. Noticeably, high-dose TBBPA-BDBPE-treated mice had fewer offspring with a female-biased sex ratio. All results show that long-term exposure to TBBPA-BDBPE caused severe reproductive impairment, including poor fertility at late reproductive age. It is therefore concluded that slight testicular injuries in early life can contribute to reproductive impairment at late reproductive age, highlighting that alterations in certain non-conventional endpoints should be noticed as well as conventional endpoints in future reproductive toxicity studies.

Effects of different grains on bacterial diversity and enzyme activity associated with digestion of starch in the foal stomach.

BACKGROUND: Compared with the stomach of ruminant cattle, the stomach of horse is small and mainly for chemical digestion, but the microorganisms in the stomach play an important role in maintaining the homeostasis of the internal environment. Due to the complexity of the microbes in the stomach, little is known about the diversity and structure of bacteria in the equine stomach. Grains are the main energy source for plant-eating livestock and energy is derived through enzymatic hydrolysis of grains into glucose or their microbial fermentation into Volatile fatty acids (VFA). However, the mechanism through which these ingested grains are chemically digested as well as the effect of these grains on the stomach remains elusive. This study explored the effects of feeding different grains (corn, oats, and barley) on bacterial diversity, structure, and composition in the foal's stomach content. Furthermore, the effects of different grains on the vitality of starch digestion-related stomach enzymes were investigated. RESULTS: No significant differences were observed (P > 0.05) in the bacterial rarefaction curves of Operational Taxonomic Units (OTUs) and diversity of the stomach microbiota in all foals. This study also revealed the statistical differences for Firmicutes, Cyanobacteria, Actinobacteria, Fibrobacteres, Lactobacillaceae, Streptococcaceae, Unidentified_Clostridiales, Prevotellaceae, Lactobacillus, Streptococcus, Unidentified_Cyanobacteria, Unidentified_Clostridiales, Lactococcus, Sphingomonas, Lactobacillus_hayakitensis, Lactobacillus_equigenerosi, and Clostridium_perfringens. The linear discriminant analysis effect size analysis revealed 9 bacteria at each classification level. The functional analysis of species information by using FAPROTAX software was able to predict 35 functions, and the top 5 functions were chemoheterotrophy, fermentation, animal_parasites_or_symbionts, nitrate_reduction, and aerobic_chemoheterotrophy. The study also revealed statistical differences for pH, glucose concentration, ß-amylase, maltase, and amylase. CONCLUSIONS: The different grains had no significant effect on the microbial diversity of the stomach content of the foal. However, the relative bacterial abundances differed significantly in response to different diets. Particularly, oats fed to the foals significantly increased the relative abundance of Firmicutes, Lactobacillaceae, Lactobacillus, and Lactobacillus_hayakitensis. The grain had no significant effect on the pH of the stomach content, glucose concentration, and enzyme viability in the foal.

Combining Precursor and Fragment Information for Improved Detection of Differential Abundance in Data Independent Acquisition.

In bottom-up, label-free discovery proteomics, biological samples are acquired in a data-dependent (DDA) or data-independent (DIA) manner, with peptide signals recorded in an intact (MS1) and fragmented (MS2) form. While DDA has only the MS1 space for quantification, DIA contains both MS1 and MS2 at high quantitative quality. DIA profiles of complex biological matrices such as tissues or cells can contain quantitative interferences, and the interferences at the MS1 and the MS2 signals are often independent. When comparing biological conditions, the interferences can compromise the detection of differential peptide or protein abundance and lead to false positive or false negative conclusions.We hypothesized that the combined use of MS1 and MS2 quantitative signals could improve our ability to detect differentially abundant proteins. Therefore, we developed a statistical procedure incorporating both MS1 and MS2 quantitative information of DIA. We benchmarked the performance of the MS1-MS2-combined method to the individual use of MS1 or MS2 in DIA using four previously published controlled mixtures, as well as in two previously unpublished controlled mixtures. In the majority of the comparisons, the combined method outperformed the individual use of MS1 or MS2. This was particularly true for comparisons with low fold changes, few replicates, and situations where MS1 and MS2 were of similar quality. When applied to a previously unpublished investigation of lung cancer, the MS1-MS2-combined method increased the coverage of known activated pathways.Since recent technological developments continue to increase the quality of MS1 signals (e.g. using the BoxCar scan mode for Orbitrap instruments), the combination of the MS1 and MS2 information has a high potential for future statistical analysis of DIA data.

Effects of steam-flaked grains on foals' growth and faecal microbiota.

BACKGROUND: There is little objective information concerning the effect of steam-flaked grains on foal's growth performance and faecal microbiota. To determine the effects of steam-flaked grains on foal's growth performance and faecal microbiota, faecal samples were collection from 18 foals which had been fed either corn, oat or barley diets over the 60 days of the experiment. Body weight and conformation measurements were collected. Next-generation sequencing of the V3 + V4 region of the 16 S rRNA gene was used to assess the microbial composition of faeces. Alpha diversity, Venn graph, Relative abundance and beta diversity are presented. RESULTS: There was a significantly higher larger increase in the body weight of those foals fed barley compared to either corn or oats. There were also significant changes in the Alpha diversity of the gut microbiota. The Shannon and Simpson indices were significantly higher in the barley fed group than those fed corn or oats. The Chao1 index was significantly higher in the oat fed group than the corn or barley fed groups. There were significant changes in the relative abundance of bacteria in the microbiota in terms of phylum, family and genus. The histogram of LDA value distribution showed that the 12 statistically different biomarkers of the bacteria were present. Tax4Fun function annotation clustering heat map showed that functional information was detected from 26 species of bacteria in faecal samples from the foals. CONCLUSIONS: Differences by starch sources were found in overall growth of the foals and in the faecal microbiota if either supplementary corn, oat or barley was fed. Further studies are required to determine the potential impact of the changes in the microbiota on the health and development of foals fed cereal starch of different sources.

Dietary administration of resistant starch improved caecal barrier function by enhancing intestinal morphology and modulating microbiota composition in meat duck.

Resistant starch (RS) was recently approved to exert a powerful influence on gut health, but the effect of RS on the caecal barrier function in meat ducks has not been well defined. Thus, the effect of raw potato starch (RPS), a widely adopted RS material, on microbial composition and barrier function of caecum for meat ducks was determined. A total of 360 Cherry Valley male ducks of 1-d-old were randomly divided and fed diets with 0 (control), 12, or 24 % RPS for 35 d. Diets supplemented with RPS significantly elevated villus height and villus height:crypt depth ratio in the caecum. The 16S rRNA sequence analysis indicated that the diet with 12 % RPS had a higher relative abundance of Firmicutes and the butyrate-producing bacteria Faecalibacterium, Subdoligranulum, and Erysipelatoclostridium were enriched in all diets. Lactobacillus and Bifidobacterium were significantly increased in the 24 % RPS diet v. the control diet. When compared with the control diet, the diet with 12 % RPS was also found to notably increase acetate, propionate and butyrate contents and up-regulated barrier-related genes including claudin-1, zonula occludens-1, mucin-2 and proglucagon in the caecum. Furthermore, the addition of 12 % RPS significantly reduced plasma TNF-α, IL-1ß and endotoxin concentrations. These data revealed that diets supplemented with 12 % RPS partially improved caecal barrier function in meat ducks by enhancing intestinal morphology and barrier markers expression, modulating the microbiota composition and attenuating inflammatory markers.

Estimating Arctic Sea Ice Thickness with CryoSat-2 Altimetry Data Using the Least Squares Adjustment Method.

Satellite altimeters can be used to derive long-term and large-scale sea ice thickness changes. Sea ice thickness retrieval is based on measurements of freeboard, and the conversion of freeboard to thickness requires knowledge of the snow depth and snow, sea ice, and sea water densities. However, these parameters are difficult to be observed concurrently with altimeter measurements. The uncertainties in these parameters inevitably cause uncertainties in sea ice thickness estimations. This paper introduces a new method based on least squares adjustment (LSA) to estimate Arctic sea ice thickness with CryoSat-2 measurements. A model between the sea ice freeboard and thickness is established within a 5 km × 5 km grid, and the model coefficients and sea ice thickness are calculated using the LSA method. Based on the newly developed method, we are able to derive estimates of the Arctic sea ice thickness for 2010 through 2019 using CryoSat-2 altimetry data. Spatial and temporal variations of the Arctic sea ice thickness are analyzed, and comparisons between sea ice thickness estimates using the LSA method and three CryoSat-2 sea ice thickness products (Alfred Wegener Institute (AWI), Centre for Polar Observation and Modelling (CPOM), and NASA Goddard Space Flight Centre (GSFC)) are performed for the 2018-2019 Arctic sea ice growth season. The overall differences of sea ice thickness estimated in this study between AWI, CPOM, and GSFC are 0.025 ± 0.640 m, 0.143 ± 0.640 m, and -0.274 ± 0.628 m, respectively. Large differences between the LSA and three products tend to appear in areas covered with thin ice due to the limited accuracy of CryoSat-2 over thin ice. Spatiotemporally coincident Operation IceBridge (OIB) thickness values are also used for validation. Good agreement with a difference of 0.065 ± 0.187 m is found between our estimates and the OIB results.

Effects of maternal dietary vitamin E on the egg characteristics, hatchability and offspring quality of prolonged storage eggs of broiler breeder hens.

This research aims to evaluate the effects of maternal vitamin E (VE) dietary supplementation on the egg characteristics, hatchability and antioxidant status of the embryo and newly hatched chicks of prolonged storage eggs. A total of 576 75-week-old Ross 308 breeder hens were randomly allocated into three dietary VE treatments (100, 200 and 400 mg/kg) with 6 replicates of 32 hens, for a 12-week feeding trial. At week 12, a total of 710 eggs were collected over a 5-day period, and eggs per treatment were attributed into 5 replicates and stored for 14 days until incubation. The egg yolk, trunk and head of 7-day-old embryo and the serum, liver, brain and yolk sac of newly hatched chicks were sampled for the evaluation of antioxidant status. Results showed that as maternal dietary VE levels increased, yolk α-tocopherol concentration increased (p < .05). Compared with 100 mg/kg VE, the use of 200 and 400 mg/kg VE increased the hatchability of set/fertile eggs and total antioxidant capacity (T-AOC) of liver and serum in chicks (p < .05), and decreased both the early embryonic mortality and the malondialdehyde (MDA) content of trunk and head in 7-day-old embryos (p < .05); moreover, 400 mg/kg VE increased the yolk T-AOC (p < .05) and decreased yolk and brain MDA content of chicks (p < .05). Brain T-AOC of chicks in 200 mg/kg VE group was improved compared to that of chicks in 100 mg/kg VE group (p < .05). In conclusion, maternal dietary VE at 200 or 400 mg/kg could increase hatchability by decreasing early embryonic mortality and increasing the antioxidant status of egg yolk, embryo and newly hatched chicks as breeder egg storage was prolonged to 14-18 days. The suitable VE level for the broiler breeder diet was 200 mg/kg as breeder egg storage was prolonged.

Dietary supplement of essential oil from oregano affects growth performance, nutrient utilization, intestinal morphology and antioxidant ability in Pekin ducks.

A study was conducted to investigate the effects of oregano essential oil (EO) on growth performance, nutrients utilization, intestinal morphology, intestinal barrier-related gene expression and antioxidant capability in meat ducks. A total of 360 1-day-old ducks were divided into three groups (12 replicates pens per diet of 10 ducks in each pen): negative control (no essential oil or antibiotic), positive control (antibiotic: 500 mg/kg aureomycin of diet) and oregano EO (100 mg/kg of diet). The experiment was carried out for 35 days. Ducks were given feed and water ad libitum. Ducks fed EO supplement showed similar body weight and feed to gain ratio to antibiotic fed ducks. EO supplementation significantly increased (p < .05) feed intake (day 1-35), jejunal villus height (VH) to crypt depth (CD) ratio, serum superoxide dismutase activities (SOD) and jejunal total antioxidant capacity (T-AOC) of ducks compared to controls. Ducks fed diets supplemented with oregano EO also had decreased (p < .05) jejunal CD, serum and hepatic malondialdehyde (MDA) concentration, and the mRNA expression of jejunal zonula occludens-3 (ZO-3) and secretory immunoglobulin A (sIgA) genes in comparison to the control group. Compared to the antibiotic supplementation group, the mRNA expression of claudin1 (CLND1) and CLND2 significantly increased (p < .05), but the mRNA expression of ZO-3 and mucin 2 markedly decreased (p < .05) in the jejunum of ducks in oregano EO supplementation group. These results suggest that oregano EO improves the antioxidant capacity and intestinal defence and structural measures and may aide in helping to maintain enteric health in production without growth-promoting antibiotics.

Range of Pulmonary Autograft Responses to Systemic Pressure Immediately After Ross Procedure.

BACKGROUND: Pulmonary autograft dilatation after Ross operation often necessitates reoperation. To understand autograft remodeling, a biomechanical understanding of human autografts after exposure to systemic pressure is required. We previously developed an ex vivo human pulmonary autograft finite element (FE) model to predict wall stress after exposure to systemic pressure. However, autograft material properties vary significantly among individuals. Our study aim was to quantify range of wall stress changes in a human autograft after Ross operation prior to remodeling based upon normal variation in human autograft mechanical properties. METHODS: A normal human autograft FE model was loaded to pulmonary and systemic arterial pressures. Stress-strain data of normal human autografts (n=24) were incorporated into an Ogden hyper-elastic model to describe autograft mechanical behavior. Autograft wall stresses at pulmonary vs. systemic pressures were examined. Autograft volume-based stress analysis was performed, based on percentage of autograft element volume exceeding 1 standard deviation (SD) above group mean stress at systemic systole. RESULTS: Mean first principal wall stresses (FPS) at systole of systemic versus pulmonary pressures were 129.29±17.47kPa versus 24.42±3.85kPa (p<0.001) at the annulus, 187.53±20.06kPa versus 35.98±2.15kPa at sinuses (p<0.001), and 268.68±23.40kPa versus 50.15±5.90kPa (p<0.001) at sinotubuluar junction (STJ). The percentage of autograft element volume that exceeded one SD above the group mean was 14.3±5.6% for FPS and 12.6±10.1% for second principal stresses. CONCLUSION: We quantified normal human autograft biomechanical responses to systemic pressure based on patient-specific material properties. Regions of peak stresses were observed in autograft sinuses and STJ regions, which corresponded clinically to locations of autograft dilation. Our results provide valuable information on predicting variations in patient-specific ex vivo FE models when population-based material properties are used in settings where patient-specific properties are unknown.

Optimization of Experimental Parameters in Data-Independent Mass Spectrometry Significantly Increases Depth and Reproducibility of Results.

Comprehensive, reproducible and precise analysis of large sample cohorts is one of the key objectives of quantitative proteomics. Here, we present an implementation of data-independent acquisition using its parallel acquisition nature that surpasses the limitation of serial MS2 acquisition of data-dependent acquisition on a quadrupole ultra-high field Orbitrap mass spectrometer. In deep single shot data-independent acquisition, we identified and quantified 6,383 proteins in human cell lines using 2-or-more peptides/protein and over 7100 proteins when including the 717 proteins that were identified on the basis of a single peptide sequence. 7739 proteins were identified in mouse tissues using 2-or-more peptides/protein and 8121 when including the 382 proteins that were identified based on a single peptide sequence. Missing values for proteins were within 0.3 to 2.1% and median coefficients of variation of 4.7 to 6.2% among technical triplicates. In very complex mixtures, we could quantify 10,780 proteins and 12,192 proteins when including the 1412 proteins that were identified based on a single peptide sequence. Using this optimized DIA, we investigated large-protein networks before and after the critical period for whisker experience-induced synaptic strength in the murine somatosensory cortex 1-barrel field. This work shows that parallel mass spectrometry enables proteome profiling for discovery with high coverage, reproducibility, precision and scalability.

WITHDRAWN: Heralds of parallel MS: Data-independent acquisition surpassing sequential identification of data dependent acquisition in proteomics.

This article has been withdrawn by the authors. This article did not comply with the editorial guidelines of MCP. Specifically, single peptide based protein identifications of 9-19% were included in the analysis and discussed in the results and conclusions. We wish to withdraw this article and resubmit a clarified, corrected manuscript for review.

Relative bioavailability of humate-manganese complex for broilers fed a corn-soya bean meal diet.

The experiment was conducted to investigate the bioavailability of manganese (Mn) from humate-Mn complex relative to Mn sulphate for the starter broilers fed a conventional corn-soya bean meal diet. A total of 560 1-day-old Arbor Acres male broiler chicks were randomly allotted to one of eight replicate cages (10 chicks per cage) for each of seven treatments in a completely randomized design involving a 2 × 3 factorial arrangement of treatments with two Mn sources (humate-Mn and Mn sulphate) and three levels of added Mn (60, 120 or 180 mg Mn/kg) plus a Mn-unsupplemented control diet containing 27.23 mg Mn/kg by analysis. At 14 days of age, the blood, liver, heart and tibia were collected for Mn analyses, and the activity and mRNA abundance of heart manganese superoxide dismutase (MnSOD). The results showed that humate-Mn supplementation decreased feed intake from day 1 to day 14, whereas it did not influence (p > 0.20) body weight at day 14 as compared to Mn sulphate. The Mn source did not influence Mn concentration in the liver, heart and tibia, and the activity and mRNA abundance of heart MnSOD, while humate-Mn decreased plasma Mn as compared to Mn sulphate. The Mn concentration in the plasma and heart, and the activity and mRNA abundance of heart MnSOD increased linearly as dietary Mn concentration increased. Based on slope ratios from multiple linear regressions of Mn concentrations in the plasma and heart, and the activity and mRNA abundance of heart MnSOD on daily intake amount of dietary analysed Mn, the bioavailability of humate-Mn complex relative to Mn sulphate (100%) was 82.8, 90.4, 82.8 and 81.9 respectively. These results indicated that the Mn from humate-Mn complex was just as bioavailable as the Mn from Mn sulphate for the starter broilers (day 1-14).

A Cost-Effective High-Throughput Plasma and Serum Proteomics Workflow Enables Mapping of the Molecular Impact of Total Pancreatectomy with Islet Autotransplantation.

Blood is an ideal body fluid for the discovery or monitoring of diagnostic and prognostic protein biomarkers. However, discovering robust biomarkers requires the analysis of large numbers of samples to appropriately represent interindividual variability. To address this analytical challenge, we established a high-throughput and cost-effective proteomics workflow for accurate and comprehensive proteomics at an analytical depth applicable for clinical studies. For validation, we processed 1 µL each from 62 plasma samples in 96-well plates and analyzed the product by quantitative data-independent acquisition liquid chromatography/mass spectrometry; the data were queried using feature quantification with Spectronaut. To show the applicability of our workflow to serum, we analyzed a unique set of samples from 48 chronic pancreatitis patients, pre and post total pancreatectomy with islet autotransplantation (TPIAT) surgery. We identified 16 serum proteins with statistically significant abundance alterations, which represent a molecular signature distinct from that of chronic pancreatitis. In summary, we established a cost-efficient high-throughput workflow for comprehensive proteomics using PVDF-membrane-based digestion that is robust, automatable, and applicable to small plasma and serum volumes, e.g., finger stick. Application of this plasma/serum proteomics workflow resulted in the first mapping of the molecular implications of TPIAT on the serum proteome.

Genome-wide DNA methylation analysis in jejunum of Sus scrofa with intrauterine growth restriction.

Intrauterine growth restriction (IUGR) may elicit a series of postnatal body developmental and metabolic diseases due to their impaired growth and development in the mammalian embryo/fetus during pregnancy. In the present study, we hypothesized that IUGR may lead to abnormally regulated DNA methylation in the intestine, causing intestinal dysfunctions. We applied reduced representation bisulfite sequencing (RRBS) technology to study the jejunum tissues from four newborn IUGR piglets and their normal body weight (NBW) littermates. The results revealed extensively regional DNA methylation changes between IUGR/NBW pairs from different gilts, affecting dozens of genes. Hiseq-based bisulfite sequencing PCR (Hiseq-BSP) was used for validations of 19 genes with epigenetic abnormality, confirming three genes (AIFM1, MTMR1, and TWIST2) in extra samples. Furthermore, integrated analysis of these 19 genes with proteome data indicated that there were three main genes (BCAP31, IRAK1, and AIFM1) interacting with important immunity- or metabolism-related proteins, which could explain the potential intestinal dysfunctions of IUGR piglets. We conclude that IUGR can lead to disparate DNA methylation in the intestine and these changes may affect several important biological processes such as cell apoptosis, cell differentiation, and immunity, which provides more clues linking IUGR and its long-term complications.

Effect of graded calcium supplementation in low-nutrient density feed on tibia composition and bone turnover in meat ducks.

Both genetic selection and increasing nutrient density for improving growth performance had inadvertently increased leg problems of meat ducks, which adversely affects animal welfare. We hypothesised that slowing weight gain with improving tibia quality probably enhanced tibial mechanical properties and alleviated leg deformities. Therefore, the present study aimed to evaluate the effect of graded Ca supplementation in a low-nutrient density (LND) diet on tibia composition and bone turnover in meat ducks. A total of 720 15-d-old male meat ducks were randomly assigned and fed a standard nutrient density positive control (PC) diet containing 0·9 % Ca, and four LND diets with 0·5, 0·7, 0·9 and 1·1 % Ca, respectively. Ducks fed the 0·5 % Ca LND diet and the PC diet had higher incidence of tibial dyschondroplasia (TD). When compared with the 0·5 % Ca LND diet, LND diets with ≥0·7 % Ca significantly improved tibia composition, microarchitecture and mechanical properties, and consequently decreased the incidence of TD. Furthermore, LND diets with ≥0·7 % Ca increased osteocyte-specific gene mRNA expression, blocked the expression of osteoblast differentiation marker genes including osteocalcin, collagenase-1 and alkaline phosphatase (ALP), and also decreased the expression of osteoclast differentiation genes, such as vacuolar-type H+-ATPase, cathepsin K and receptor activator of NF-κB. Meanwhile bone markers such as serum ALP, osteocalcin (both osteoblast markers) and tartrate-resistant acid phosphatase (an osteoclast marker) were significantly decreased in at least 0·7 % Ca treated groups. These findings indicated that LND diets with ≥0·7 % Ca decreased bone turnover, which subsequently increased tibia quality for 35-d-old meat ducks.

Impact of Patient-Specific Material Properties on Aneurysm Wall Stress: Finite Element Study.

BACKGROUND: Finite element analysis (FEA) can be used to determine ascending thoracic aortic aneurysm (aTAA) wall stress as a potential biomechanical predictor of dissection. FEA is dependent upon zero-pressure three-dimensional geometry, patient-specific material properties, wall thickness, and hemodynamic loading conditions. Unfortunately, determining material properties on unoperated patients using non-invasive means is challenging; and we have previously demonstrated significant material property differences among aTAA patients. Our study objective was to determine the impact of patient-specific material properties on aTAA wall stress. Using FEA, we investigated if patient-specific wall stress could be reasonably predicted using population-averaged material properties, which would greatly simplify dissection prediction. METHODS: ATAA patients (n=15) with both computed tomography (CT) imaging and surgical aTAA specimens were recruited. Patient-specific aTAA CT geometries were meshed and pre-stress geometries determined as previously described. Patient-specific material properties were derived from biaxial stretch testing of aTAA tissue and incorporated into a fiber-enforced hyper-elastic model, while group-averaged material properties were estimated using mean values of each parameter. Population-averaged material properties were also calculated from literature and studied. Wall stress distribution and its magnitude were determined using LS-DYNA FEA software. Peak and averaged stresses and stress distributions were compared between patient-specific and both group- and population-averaged material property models. RESULTS: Patient-specific material properties had minimal influence on either peak or averaged wall stress compared to use of group- or population-averaged material properties. Stress distribution was also nearly superimposed among models with patient-specific vs. group- or population-averaged material properties and provided similar prediction of sites most prone to rupture. CONCLUSIONS: FEA using population-averaged material properties likely provides reliable stress prediction to indicate sites most prone to rupture. Population-averaged material properties may be reliably used in computational models to assess wall stress and significantly simplify risk prediction of aTAA dissection.